Understanding gene function is a fundamental but still challenging task. It often involves monitoring gene transcription and regulation in both live cells and tissues in real time with reporter gene assays. A reporter gene that can work in both live cells and animals would be very valuable, but is currently unavailable. The overall goals of this research are to develop an integrated reporter gene system that is suitable for imaging gene expression in both single living cells and whole living animals including humans. An ampicillin resistance gene (encoding a bacterial enzyme, beta-lactamase) is an outstanding reporter for optical measurement of gene expression in single living mammalian tissue culture cells. To extend this reporter gene to whole living animals, it will require the development of new substrates. We will design and synthesize new substrates for imaging transgene expression with positron emission tomography (PET) in whole living mice. These new probes will be validated in living mice with tumor xenografts. Their sensitivity and specificity will be assessed and compared to the existing PET imaging reporter gene assays. This work will provide a unified reporter gene system that can monitor gene expression from single live cells undergoing high-throughput selection to whole living animals with PET. Such a versatile system should find wide applications in biology from in vivo cell trafficking, tumor metastasis, small animal disease models, transgenic mice, drug screenings in mice, to in vivo gene function and regulation study.